10x Genomics Chromium

10x Genomics Chromium Controller – High-throughput automated barcoding and library construction for powerful new RNA and DNA sequencing applications. The Chromium Controller is powered by 10x GemCode Technology and enables the encapsulation in a single run of up to 80,000+ individual cells or from as little as 1 ng of HMW gDNA into 100,000s to 1,000,000s of uniquely barcoded picoliter droplets for downstream genomic analysis with Chromium Software Suite. Chromium technical applications have revolutionized single-cell RNA-seq and 10x Linked-Read technology now provides long-range information from short-read sequencing data.

Chromium solutions include:

A list of publications using 10x technology is available here. Additional educational resources including seminars, software demos, and recorded workshops can be viewed here. Datasets are also accessible from here at your convenience.

10x Genomics develop a suite of tools to analyze the DNA and RNA data produced by the Chromium controller.

  • Bclprocessor:  Generates special type of fastq files.
  • Long Ranger: Generates high-quality haplotype phasing and structural variant calls.
  • Supernova: De novo Genome assembly
  • Loupe: Desktop genome browser for data visualization
  • Loupe Cell Browser: Visualize and analyze 10x Chromium™ Single Cell 5′ and 3′ gene expression data
  • Loupe™ V(D)J Browser: Analyze, search, and visualize V(D)J sequences and clonotypes from single cell 5′ data produced by the 10x Chromium Platform.
Consultation and Assistance

Contact for Genomics and Bioinformatics Consultation
Please use this link to send an email to Dr. Magdy Alabady for consultation on new or existing Genomics and Bioinformatics projects. Also, you can contact Dr. Alabady for assistance with grant proposals and for obtaining a letter of support from GGBC.

Contact for 10x Genomics Technical Assistance
For technical questions about existing or new 10x Genomics projects, please contact the following Staff:

Sample Preparation

Sample Preparation for Chromium Linked-reads

For the genomics application, high quality HMW DNA is required. DNA will be tested on the Fragment Analyzer using the 50Kb protocol and, if needed, on the PippinPuls (a PFGE system).

10X Genomics recommendations for specific HMW DNA isolation protocols can be accessed from here.

  • DNA should be above 50kb with greater than 80% of the DNA at 100kb or larger. After isolation, the DNA should be handled very gently (minimal pipetting, use of wide-bore tips, avoid freeze/thaw, etc).
  • Ship HMW DNA on dry ice for stability, but once it arrives at the GSL it will be stored at 4C.
  • Though the protocol uses 1ng of HMW DNA as input, a minimum of 500ng of DNA in 20ul of water or Tris should be submitted to allow for QC and visualization of the HMW DNA on a gel.
  • If you are are submitting 24 or more samples please submit samples in a 96-well plate and include an Excel file with the sample layout in your order.
  • PLEASE USE V BOTTOM PLATES AND NOT ROUND/FLAT BOTTOM PLATES.

 

Sample Preparation for Chromium Single Cell and nuclei RNAseq

  • For pricing please see our website at the link here for 10X library prep and the link here for NextSeq runs. Official quotes can be provided upon request.
  • We can process up to 8 samples on a 10X chip at a time. The library prep cost is reflective of that and cost per sample is lowest for 8 samples. We follow 10X’s sequencing recommendations, which you can view at the link here. Read lengths can be changed as desired, but please note that shorter transcript reads can reduce alignment rates. Generally a minimum of 20,000 read pairs per cell is needed. The type and number of sequencing run(s) depends on the number of samples and coverage you need. We can make libraries with between 500 and 10,000 cells per sample, though the recommended starting point is 1,600 cells the first time you do the experiment. It is recommended to try the protocol with just one sample first before moving forward with a larger number.
  • 10X has a list of demonstrated cell prep protocols at the link here. Customers should work with the tissue in their lab and provide us with a prepared cell suspension. Cell preparation methods can vary between different tissue types. Please refer to 10X’s recommendations and check the literature to find methods appropriate for your sample type. Cells are preferable in most cases, but nuclei can be used as well. Feel free to reach out to 10X or GGBC for further recommendations.
  • When you submit your order and have the cell suspension ready for library prep we need an accurate measure of cell concentration and viability. These QC data are critical to the protocol working correctly. The ideal concentration is between 700-1,200 cells/uL and the viability should be >90%. An inaccurate concentration could clog the chip or result in low yield, and the presence of dead cells will reduce the recovery rate and result in a high level of noise in the data. It is also important to minimize the time from cell prep to loading the 10X chip.
  • We do not have the equipment for cell counting and viability here at GGBC, so we have to rely on your data. We’ve seen the best results from customers who prep their cells, sort and count by flow cytometry, and then bring them on ice immediately to our lab. There is a flow cytometer in another core lab on campus that you can use. We’ve also had good results from a few customers who did the final steps of cell prep in our lab before counting and assessing viability by microscopy immediately prior to loading the 10X chip. If you want to go that route you would need to bring a scope and whatever other materials you need.
  • Please contact Julia Portocarrero at least 2 weeks in advance before submitting a single cell order.
Chromium Data and Sequencing

Chromium libraries have a unique structure that requires special sequencing considerations. Each DNA sample is indexed with four i7 barcodes sequenced as a standard i7 index read of 8bp in length. Each i7 barcode is a combination of 8-bases unique barcodes. The unique barcode used to link reads is found at the beginning of read 1 and is 16bp in length.

Chromium Linked-reads

  • 10X Genomics recommend 30-40X for whole genome resequencing projects and 40-56X coverage for de novo assembly of genomes.
  • At GGBC, we typically sequence 10X Genomic library on a NextSeq PE150 High Output run. The yield of this run is about 110-120b and 85% of the bases are >= Q30. Because the i7 barcodes are unique, two 10X libraries can be multiplexed and sequenced on the same flowcell.

Chromium Single Cell and nuclei RNAseq

  • <Coming Soon>
Prices and Quotes

Contact for Financial Inquiries and Quote Requests

Please email Kim and Elizabeth at ggbc@uga.edu, for financial inquiries or to request a quote. Be as specific as possible, so that they can more quickly assist you.

 

Genomics Linked Reads

Table 1. 10x Genomics linked-reads library preparation fees (prices valid for libraries prepared AND sequenced at GGBC ONLY*)

Number of Sample(s)UGA FeeNon-UGA FeeCommercial Fee
1$609.00$719.00$762.00
2$955.00$1,127.00$1,194.00
3$1,301.00$1,536.00$1,627.00
4$1,648.00$1,945.00$2,060.00
5$2,208.00
$2,606.00$2,760.00
6$2,555.00$3,015.00$3,194.00
7$2,901.00$3,423.00$3,627.00
8$3,247.00$3,832.00$4,059.00

* Prices are significantly higher If libraries are to be prepared at GGBC but sequenced elsewhere. Please inquire for pricing.

Table 2. Suggested Illumina sequencing run type and associated fees for linked-reads libraries

Service Description(s)UGA FeeNon-UGA FeeCommercial Fee
Library Pooling up to 2-24 samples by qPCR*$129.00$153.00$194.00
Pre-Sequencing QC (Qubit, FA, Kapa)$120.00$142.00$180.00
NextSeq (300 Cycles) Mid Output flow cell $2,383.00**$2,812.00$3,575.00
NextSeq (300 Cycles) High Output flow cell$5,548.00**$6,547.00$8,322.00
Single Cell/Nuclei RNA seq

Table 3. 10x Genomics single cell/nuclei RNA seq library preparation fees (prices valid for libraries prepared AND sequenced at GGBC ONLY*)

Number of Sample(s)UGA FeeNon-UGA FeeCommercial Fee
1$1,731.00$2,043.00$2,164.00
2$3,151.00$3,719.00$3,939.00
3$4,571.00$5,394.00$5,714.00
4$5,990.00$7,069.00$7,488.00
5$7,623.00
$8,996.00$9,529.00
6$9,042.00$10,670.00$11,303.00
7$10,462.00$12,346.00$13,078.00
8$11,882.00$14,021.00$14,853.00

* Prices are significantly higher If libraries are to be prepared at GGBC but sequenced elsewhere. Please inquire for pricing.

Table 4. Suggested Illumina sequencing run type and associated fees for single cell/nuclei RNAseq libraries

Service Description(s)UGA FeeNon-UGA FeeCommercial Fee
Library Pooling up to 2-24 samples by qPCR*$129.00$153.00$194.00
Pre-Sequencing QC (Qubit, FA, Kapa)$120.00$142.00$180.00
NextSeq (150 Cycles) Mid Output flow cell $1,638.00**$1,933.00$2,457.00
NextSeq (150 Cycles) High Output flow cell***$3,621.00**$4,273.00$5,432.00
NextSeq (75 Cycles) High Output flow cell$2,093.00**$2,470.00$3,140.00
scATAC-seq

Table 5. 10x Genomics scATAC-seq library preparation fees (prices valid for libraries prepared AND sequenced at GGBC ONLY*)

Number of Sample(s)UGA FeeNon-UGA FeeCommercial Fee
1$2,124.00$2,507.00$2,655.00
2$3,690.00$4,355.00$4,613.00
3$5,256.00$6,203.00$6,570.00
4$6,823.00$8,052.00$8,529.00
5$8,602.00
$10,151.00$10,753.00
6$10,168.00$11,999.00$12,710.00
7$11,734.00$13,847.00$14,668.00
8$13,300.00$15,694.00$16,625.00

* Prices are significantly higher If libraries are to be prepared at GGBC but sequenced elsewhere. Please inquire for pricing.

Table 6. Suggested Illumina sequencing run type and associated fees for scATAC-seq libraries

Service Description(s)UGA FeeNon-UGA FeeCommercial Fee
Library Pooling up to 2-24 samples by qPCR*$129.00$153.00$194.00
Pre-Sequencing QC (Qubit, FA, Kapa)$120.00$142.00$180.00
NextSeq (150 Cycles) High Output flow cell $3,621.00**$4,273.00$5,432.00
NextSeq (75 Cycles) High Output flow cell$2,093.00**$2,470.00$3,140.00

*Only if pooling is required
** Additional 5% discount available for UGA customers doing 10 or more NextSeq runs between 7/01/19 and the end of the UGA fiscal year, 6/15/2020. Click here for more details.
***Recommended run type